ABSTRACT
<p><b>OBJECTIVE</b>To explore method of recombinant gene lentivirus containing LIM mineralization protein-1 (LMP-1) in transfecting bone marrow mesenchymal stem cells (BMSC), and to observe the effect of gene LMP-1 on proliferation effect and expression of BMSC.</p><p><b>METHODS</b>Six clean SD rats aged 4 weeks were selected, bone marrow mesenchymal stem cells were extracted under sterile conditions and cultured to the third generation, then divided into three groups:control group (the third generation of BMSC), lentiviral vector transfection group (PGC-FU-GFP and Polybrene were injected into the third generation of BMSC) and recombinant gene transfection group (PGC-FU-LMP-1-GFP and Polybrene transfection were injected into the third generation of BMSC). After 48 hours' transfection, fluorescent expression were detected under immuno-fluorescence microscopy; lentiviral transfection efficiency were detected by flow cytometry; effect of lentiviral transfection on BMSC were evaluated by MTT; gene expression of transfected cells were determined by Western Blot.</p><p><b>RESULTS</b>1) The third generation of BMSC was cultured successfully,and transfected with MOI:100. After 48 hours, green fluorescent expression were detected and transfection efficiency was 67% under immuno-fluorescence microscopy; 2) Compared to control group, there were no statistical differences between control group and other two groups; 3) Western blot teast showed that 72KDa specific band was observed in recombinant gene transfection group and its size was similar to LMP-1 fusion protein (50 kDa+28 kDa=78 kDa).</p><p><b>CONCLUSION</b>There is no effect of recombinant gene lentivirus containing LIM on BMSC, and can effectively influence the expression of LMP-1.</p>
Subject(s)
Animals , Female , Humans , Male , Rats , Adaptor Proteins, Signal Transducing , Genetics , Metabolism , Cell Proliferation , Cells, Cultured , Cytoskeletal Proteins , Genetics , Metabolism , Genetic Therapy , Genetic Vectors , Genetics , Metabolism , LIM Domain Proteins , Genetics , Metabolism , Lentivirus , Genetics , Metabolism , Mesenchymal Stem Cells , Cell Biology , Metabolism , Virology , Osteoporosis , Genetics , Therapeutics , Rats, Sprague-DawleyABSTRACT
<p><b>BACKGROUND</b>The failure of hormone treatment for advanced prostate cancer might be related to aberrant activation of the androgen receptor. We have shown that (125)I labeled triple-helix forming oligonucleotide (TFO) against the androgen receptor gene inhibits androgen receptor expression and cell proliferation of LNCaP prostate cancer cells in vitro. This study aimed at exploring the effects of the (125)I-TFO on prostate tumor growth in vivo using a nude mouse xenograft model.</p><p><b>METHODS</b>TFO was labeled with (125)I by the iodogen method. Thirty-two nude mice bearing LNCaP xenograft tumors were randomized into 4 groups and were intratumorally injected with (125)I-TFO, unlabeled TFO, Na(125)I and normal saline. Tumor size was measured weekly. The tumor growth inhibition rate (RI) was calculated by measurement of tumor weight. The expression of the androgen receptor gene was performed by RT-PCR and immunohistochemical study. The prostate specific antigen (PSA) serum levels were measured by enzyme linked immunosorbent assay. The tumor cell apoptosis index (AI) was detected by TUNEL assay.</p><p><b>RESULTS</b>Tumor measurements showed that tumor development was significantly inhibited by either (125)I-TFO or TFO, with tumor RIs of 50.79% and 32.80% respectively. (125)I-TFO caused greater inhibition of androgen receptor expression and higher AIs in tumor tissue than TFO. Both the tumor weight and the PSA serum levels in (125)I-TFO treated mice ((0.93 +/- 0.15) g and (17.43 +/- 1.85) ng/ml, respectively) were significantly lower than those ((1.27 +/- 0.21) g and (28.25 +/- 3.41) ng/ml, respectively) in TFO treated mice (all P < 0.05). Na(125)I did not significantly affect tumor growth and androgen receptor expression in tumor tissue.</p><p><b>CONCLUSIONS</b>The (125)I-TFO can effectively inhibit androgen receptor expression and tumor growth of human prostate cancer xenografts in vivo. The inhibitory efficacy of (125)I-TFO is more potent than that of TFO, providing a reference for future studies of antigen radiotherapy.</p>
Subject(s)
Animals , Humans , Male , Mice , Androgen Receptor Antagonists , Cell Line, Tumor , Cell Proliferation , Enzyme-Linked Immunosorbent Assay , Immunohistochemistry , Iodine Radioisotopes , Mice, Inbred BALB C , Mice, Nude , Oligonucleotides , Chemistry , Pharmacology , Therapeutic Uses , Prostate-Specific Antigen , Blood , Prostatic Neoplasms , Drug Therapy , Metabolism , Pathology , Receptors, Androgen , Genetics , Metabolism , Reverse Transcriptase Polymerase Chain Reaction , Tumor Burden , Xenograft Model Antitumor Assays , MethodsABSTRACT
<p><b>OBJECTIVE</b>To investigate the effect of antibody-targeted chemotherapy against human prostate cancer LNCaP cells in vitro.</p><p><b>METHODS</b>The monoclonal antibody 7E11C5.3 against human prostate cancer was conjugated to pingyangmycin (PYM), mediated by dextran T-40, and the immunoreactivity of 7E11C5.3 was determined by indirect enzyme-linked immunosorbent assay. The bacteriostatic activity of the conjugate was determined using TTC assay, and its cytotoxicity against LNCaP cells was determined by MTT assay.</p><p><b>RESULTS</b>The 7E11C5.3:PYM molar ratio was l:54 in the conjugate, and the immunoreactivity of 7E11C5.3 was decreased by approximately 10% to 20% after conjugation. The bacteriostatic activity of conjugated PYM was 25% of that of free PYM. The 50% inhibitory doses (IC50) of 7E11C5.3-PYM conjugate and free PYM against the in vitro cultured LNCaP cells were 9.41-/+1.98 microg/ml and 29.92-/+7.88 microg/ml, respectively.</p><p><b>CONCLUSION</b>7E11C5.3-PYM conjugate displays stronger cytotoxicity against anti-prostate cancer effects than free PYM.</p>